7KOV

Crystal structure of Azotobacter vinelandii 3-mercaptopropionic acid dioxygenase in complex with thiocyanate

Summary for 7KOV

• Entry DOI: 10.2210/pdb7kov/pdb
• Related: 6XB9
• Descriptor: Cysteine dioxygenase type I protein, FE (III) ION, THIOCYANATE ION, ... (5 entities in total)
• Functional Keywords: non-heme iron, facial triad, oxidoreductase, thiocyanate
• Biological source: Azotobacter vinelandii
• Total number of polymer chains: 4
• Total formula weight: 94171.15

Authors

Kiser, P.D. (deposition date: 2020-11-10, release date: 2021-03-03, Last modification date: 2023-10-18)

Primary citation

York, N.J.,Lockart, M.M.,Sardar, S.,Khadka, N.,Shi, W.,Stenkamp, R.E.,Zhang, J.,Kiser, P.D.,Pierce, B.S.
Structure of 3-mercaptopropionic acid dioxygenase with a substrate analog reveals bidentate substrate binding at the iron center.
J.Biol.Chem., 296:100492-100492, 2021

PubMed Abstract: Thiol dioxygenases are a subset of nonheme iron oxygenases that catalyze the formation of sulfinic acids from sulfhydryl-containing substrates and dioxygen. Among this class, cysteine dioxygenases (CDOs) and 3-mercaptopropionic acid dioxygenases (3MDOs) are the best characterized, and the mode of substrate binding for CDOs is well understood. However, the manner in which 3-mercaptopropionic acid (3MPA) coordinates to the nonheme iron site in 3MDO remains a matter of debate. A model for bidentate 3MPA coordination at the 3MDO Fe-site has been proposed on the basis of computational docking, whereas steady-state kinetics and EPR spectroscopic measurements suggest a thiolate-only coordination of the substrate. To address this gap in knowledge, we determined the structure of Azobacter vinelandii 3MDO (Av3MDO) in complex with the substrate analog and competitive inhibitor, 3-hydroxypropionic acid (3HPA). The structure together with DFT computational modeling demonstrates that 3HPA and 3MPA associate with iron as chelate complexes with the substrate-carboxylate group forming an additional interaction with Arg168 and the thiol bound at the same position as in CDO. A chloride ligand was bound to iron in the coordination site assigned as the O-binding site. Supporting HYSCORE spectroscopic experiments were performed on the (3MPA/NO)-bound Av3MDO iron nitrosyl (S = 3/2) site. In combination with spectroscopic simulations and optimized DFT models, this work provides an experimentally verified model of the Av3MDO enzyme-substrate complex, effectively resolving a debate in the literature regarding the preferred substrate-binding denticity. These results elegantly explain the observed 3MDO substrate specificity, but leave unanswered questions regarding the mechanism of substrate-gated reactivity with dioxygen.

PubMed: 33662397
DOI: 10.1016/j.jbc.2021.100492

Experimental method

X-RAY DIFFRACTION (2.95 Å)